Design synthetic 3D scaffold for skeletal muscle tissue differentiation

Skeletal muscle (SKM) loss is largely irreversible, and current therapeutic options to restore muscle mass remain limited. Despite the regenerative capacity of SKM due to resident satellite cells, in vitro and in vivo studies using stem cells have shown low regenerative and self-renewal properties. Conventional 2D monolayer culture systems fail to replicate the in vivo environment due to the lack of cell-cell and cell-matrix interactions, often resulting in misleading outcomes. New strategies are needed to address the limitations of current tissue repair and regeneration methods.Tissue engineering (TE) has emerged as a promising approach for SKM tissue renewal both in vitro and in vivo. TE combines scaffolds, cells, and regulatory signals to create a biomimetic extracellular matrix (ECM), supporting cell attachment and proliferation. This study evaluated a poly (L-lactic acid)-co-poly-(ε-caprolactone) (PLCL) electrospun scaffold coated with type I collagen to support SKM fabrication in vitro. The PLCL scaffold promoted myoblast attachment, elongation, and myotube formation, offering insights into 3D tissue-like structure development. To investigate SKM functionality, Atomic Force Microscopy (AFM) was used to monitor contractile activity on the scaffold, simulating excitation-contraction coupling. A 3D bioprinted scaffold with collagen, laminin, and fibronectin mimicked the native SKM environment. This model facilitated myoblast differentiation from healthy (CTRL) and Limb-Girdle Muscular Dystrophy Type D2 (LGMDD2) conditions, analyzing Myogenic Regulatory Factors (MRFs), MEF2C isoforms, and splicing factors like SRSF1 and RBM4, cargoes of TNPO3 implicated in LGMDD2 pathology. Using a micropillar system, early differentiation impairments caused by TNPO3 mutations were identified.These findings highlight the potential of TE-based models in understanding SKM development and LGMDD2 pathology while offering platforms for drug screening and therapeutic exploration. Future studies aim to refine these TE platforms for targeted therapeutic applications and improved muscular dystrophy treatments.